PROJECT SUMMARY Although poly (ADP-ribose) polymerase inhibitor (PARPi) has emerged as a promising drug for patients with cancer, primary and acquired resistance is a major clinical problem for PARPi in cancer treatment. Several drug combination strategies have been designed and evaluated in preclinical and early clinical trials to overcome this challenge. Therefore, strategies to enhance response to PARPi in primary and acquired homologous recombination (HR)-proficient tumors would represent a significant advance in cancer care. Bromodomains and extra-terminal domain inhibitor (BETi) has been rapidly advanced into early clinical trials and has shown impressive anti-tumor activity. Given that clinical activity of BETi alone may be insufficient to manage patients according to recent clinical trials, the combination of BETi with other treatment methods need to be designed and evaluated. Using a drug synergistic screen that combined a PARPi with 20 well-characterized epigenetic drugs, we identified BETi as a drug that acted synergistically with PARPi in HR-proficient cancer cells. Functional assays demonstrated that repressed BET activity reduces HR and subsequently enhances PARPi-induced DNA damage in cancer cells. Chemical inhibition or genetic depletion of BET proteins impairs transcription of several essential genes in HR. Moreover, BETi treatment sensitized tumors to PARP inhibition in preclinical animal models of HR-proficient breast and ovarian cancers. Finally, we showed that the BRD4 gene was significantly and focally amplified across common adult cancers, although its gene fusion was a rare genomic alteration. Thus, we hypothesize that BETi may suppress HR and enhance NHEJ, thereby sensitizing HR-proficient cancer cells to PARP inhibition. Aim 1. Characterize the molecular mechanisms by which BETi synergistically acts with PARPi. Aim 2. Evaluate the combination therapy of BET and PARP inhibitors in preclinical models. Aim 3. Define immune responses to BETi and PARPi treatment in the tumor microenvironment. Our proposed studies may provide strong rationale for clinical application of PARPi in the setting of combination with BETi to treat both cancers with de novo resistance to PARPi therapy and cancers with acquired resistance. Therefore, combination with BETi could greatly expand the utility of PARP inhibition to patients with HR-proficient cancer.